1. Field of the Invention
This invention relates to the delivery of bioactive substances. In particular, the invention relates to the delivery of effector units in animals, and to the modulation of their release.
2. Summary of the Related Art
Drugs are compounds almost always foreign to the body. The processes of inputting, distributing and eliminating drugs are therefore of paramount importance in determining the onset, duration and intensity of drug effect. Hence, in the context of drug development, the ability to modulate the rate at which a particular drug becomes systemically available, and to direct its bioavailability to a particular tissue are often of crucial significance. Pharmakokinetic, pharmacological and toxological constraints create the need to modulate drug delivery to minimize side effects or to optimize drug efficacy.
Drug delivery involves both drug absorption (the process of movement from the site of administration toward systemic circulation) and drug distribution or bioavailability (the process by which a drug becomes available at the site of action. Direct placement of a drug into the bloodstream (usually i.v.) ensures complete delivery of the dose to the general circulation only. Because drug distribution to other tissues is often delayed, direct administration may result in the accumulation of high plasma concentrations of the drug administered immediately postinfusion. Notably, high plasma concentrations (beyond the therapeutic window for a particular drug regiment) often cause a wide variety of toxic drug reactions which are dose-related. In addition, for many drugs and/or metabolites, distribution to the active site or tissue necessitate movement across biological barriers, for example via passive diffusion, facilitated diffusion or pinocytosis. For most drugs, distribution is not limited to the desired site (i.e., the site of action) thereby reducing their efficacy and increasing the chances of undesirable side effects.
Therefore, there is a need to develop delivery systems and methods suitable to control drug release following administration and to direct drug diffusion through biological barriers in a selective fashion. Gene therapy, and more specifically antisense therapy, is emblematic of a promising pharmacological approach for which there is a pressing need to control drug release and direct drug delivery to particular sites of action.
The potential for using oligonucleotides as inhibitors of specific gene expression in an antisense therapeutic approach was first suggested in the late ""70s. (Paterson et al., Proc. Natl. Acad. Sci. USA 74: 4370-4374 (1977); Zamecnik and Stephenson, Proc. Natl. Acad. Sci. USA 75: 280-284 and 285-288 (1978)). To date, the ability of antisense oligonucleotides to inhibit virus propagation has become firmly established. (See e.g., Agrawal, Trends in Biotechnology 10: 152-158 (1992). Antisense oligonucleotides have also been developed as anti-parasitic agents. (See e.g., PCT publication no. WO93/13740; Tao et al., Antisense Research and Development 5: 123-129 (1995)). More recently, antisense oligonucleotides have shown promise as candidates for therapeutic applications for diseases resulting from expression of cellular genes. (See e.g., PCT publication no. WO95/09236, PCT publication no. WO94/26887, and PCT application no. PCT/US94/13685). The development of various antisense oligonucleotides as therapeutic and diagnostic agents has recently been reviewed by Agrawal and Iyer, Current Opinion in Biotechnology 6: 12-19 (1995).
Much is currently being discovered about the pharmacodynamic properties of oligonucleotides. Agrawal et al., Clinical Pharmacokinetics 28: 7-16 (1995) and Zhang et al., Clinical Pharmacology and Therapeutics 58: 44-53 (1995). Some of these new studies have led to new challenges to be overcome for the optimization of oligonucleotides as therapeutic agents. Such optimization should include elimination or reduction of cardiovascular side effects and in vivo instability problems. Henry et al., Pharm. Res. 11: PPDM8082 (1994) discloses that oligonucleotides may potentially interfere with blood clotting. Cardiovascular side effects are believed to stem from high plasma concentrations of oligonucleotides which have been observed immediately postinfusion. Effective in vivo stability of oligonucleotides is affected by both their degradation and their elimination. With respect to in vivo stability of oligonucleotides, it has been shown that following intravenous administration to mice, rats or monkeys, oligonucleotides are degraded mainly from the 3xe2x80x2- end. (Temsamani et al., Antisense and Nucleic Acid Drug Development (in press)). Notably, because of instability considerations, higher dosages of oligonucleotides are necessary. Gailbraith et al., Antisense Research and Development 4:201-206 (1994) discloses complement activation and depletion by phosphorothioate oligonucleotides. Recently, several studies on elimination of oligonucleotides have been published. Agrawal et al., Proc. Natl. Acad. Sci. (USA) 88: 7595-7599 (1991) describes the intravenous and intraperitoneal administration of a 20 mer phosphorothioate linked-oligonucleotide to mice. In this study, approximately 30% of the administered dose was excreted in the urine over the first 24 hours with accumulation preferentially in the liver and kidney. Plasma half-lives ranged from about 1 hour (txc2xdxcex1) and 40 hours (txc2xdxcex2), respectively. Similar results have been reported in subsequent studies (Iversen, Anti-Cancer Drug Design 6:531-538(1991); Iversen, Antisense Res. Devel. 4:43-52 (1994); and Sands, Mol. Pharm. 45:932-943 (1994)).
Dehydration and elimination of many other drugs may also affect their efficacy and therapeutic window. Therefore, there remains a need to develop more effective therapeutic methods for modulating the release and direct the delivery of drugs which can be easily manipulated to fit the animal and condition to be treated while producing fewer side effects.
This invention relates to bioactive substances with enhanced pharmakokinetic properties and to their use in biomedical applications. In particular, the invention relates to substances including carriers for the delivery of effector units in animals, and for the modulation of the release of the same. The present inventor has discovered that bioreversible association of a bioactive substance with a hyperstructure forming unit allows modulated release and directed delivery of the bioactive substance. Thus, the invention provides compounds and methods for modulating the release and direct the delivery of drugs which can be easily manipulated to fit the animal and condition to be treated while producing fewer side effects.
In a first aspect, the invention provides a bioactive substance including a hyperstructure forming unit and an effector unit, wherein there is an operable association between the hyperstructure forming unit and the effector unit.
In a preferred embodiment, the bioactive substance is an oligonucleotide including the bioreversibly attached nucleic acid sequence GGGG or GTGT capable of aggregating with other oligonucleotides in such a way as to assemble hyperstructures of two or more oligonucleotides.
In a second aspect, the invention provides methods for delivering the effector units of this invention. In a preferred embodiment of this aspect, the invention provides a method for delivering an effector unit in an animal, including a human, comprising administering to the animal a therapeutically effective amount of a bioactive substance according to the present invention, for a therapeutically effective period of time.
In a third aspect, the invention provides a method for investigating the biochemical and biophysical roles of particular genes. In the method according to this aspect of the invention, a bioactive substance with a nucleic acid sequence complementary to a target sequence of interest, is introduced in the cell type of interest. The bioactive substance of this invention can be administered at different points in the cell cycle, or in conjunction with promoters or inhibitors of cell growth to determine the role of the target sequence in the growth of the cell type of interest.